The objective of this project is to study the properties of allospecific T-cell lines. Recent cloning experiments have shown that clones of one of the lines are of Lyt 1+, Lyt 2- phenotypeand are able to perform at least three different functions: They lyse spleen blast cell targets; cooperate with B cells for a humoral response to SRBC; and if injected with stimulator cells into the footpads of mice, will evoke a delayed type hypersensitivity reaction. These results show that the Lyt phenotype is not related to the function of T cells and that T cells may be polyfunctional. These results also raise the question of how killer cells may be able to cooperate with B cells for a humoral response. This appears to be possible via factors which are released from these T cells. In current experiments we are studying the properties of these factors. Since these factors are not only released from T cells upon specific allogeneic stimulation but also by transformed derivatives of these T cell lines, it is now possible to initiate large-scale preparation and purification of lymphokines. Three activities are presently being purified: TCGF, BCGF, and TRF. TCGF has been purified and is presently being sequenced, while the other two lymphokines are in various stages of purification. In another facet of this study we are studying with light microscopy the events leading to target cell lysis, i.e., rearrangement of Golgi, MTOC and granules after target cell binding. We are also studying perforin molecules secreted by killer cells, which appear to cause target lysis by channel formation.